The long-term goal of this project is to understand the regulatory mechanisms which prevent cells from proliferating though they have the capacity to do so. These mechansisms are extremely important to the organism since their failure would result in catastrophic, uncontrolled growth. Therefore, elucidation of growth control mechanisms is one of the prime objectives of cancer research. The reaction of a variety of substances, mitogens, with the lymphocyte membrane can alter metabolism and induce proliferation in this normally quiescent cell. An understanding of the essential processes which cause activation may allow one to determine how cells maintain a quiescent state. One problem in interpreting biochemical data from mitogen activation experiments is the heterogeneity of lymphocyte cultures. A sizable proportion of cells may not respond to stimulant and responsive cells are extremely asynchronous with respect to the time at which they synthesize DNA. The immediate objectives of this project are 1. to study further the behavior of lymphocytes in culture using DNA density transfer technology and 2. to develop a method of isolating homogeneous subpopulations of quiescent human lymphocytes that can be activated in a relatively synchronous manner. The strategy of the latter is based, in part, on the observation of my colleagues and me that mitogen activated lyphocytes divide and then become quiescent unless stimulant is present. Thus, activation can be studied in the quiescent progeny cells since, like their progenitors, reactivation also requires reaction of mitogen with the cell membrane. A more homogeneous population of mitogen responsive lymphocytes may be obtained using a method I propose to develop based on my previously published work. Activated lymphocytes committed to synthesize DNA remain attached to sepharose beads (which have concanavalin A covalently bound) even in the presence of the binding competitor methyl-a-d-mannoside (MAM). The MAM can dissociate unactivated or uncommitted cells from the beads. Physical separation of the dissociatable populations from the nondissociatable population followed by removal of attached cells by mechanical means should be helpful in isolating more homogeneous lymphocyte cultures for further study.